Type 3 innate lymphoid cells maintain intestinal epithelial stem cells after tissue damage

Disruption of the intestinal epithelial barrier allows bacterial translocation and predisposes to destructive inflammation. To ensure proper barrier composition, crypt-residing stem cells continuously proliferate and replenish all intestinal epithelial cells within days. As a consequence of this high mitotic activity, mucosal surfaces are frequently targeted by anticancer therapies, leading to dose-limiting side effects. The cellular mechanisms that control tissue protection and mucosal healing in response to intestinal damage remain poorly understood. Type 3 innate lymphoid cells (ILC3s) are regulators of homeostasis and tissue responses to infection at mucosal surfaces. We now demonstrate that ILC3s are required for epithelial activation and proliferation in response to small intestinal tissue damage induced by the chemotherapeutic agent methotrexate. Multiple subsets of ILC3s are activated after intestinal tissue damage, and in the absence of ILC3s, epithelial activation is lost, correlating with increased pathology and severe damage to the intestinal crypts. Using ILC3-deficient Lgr5 reporter mice, we show that maintenance of intestinal stem cells after damage is severely impaired in the absence of ILC3s or the ILC3 signature cytokine IL-22. These data unveil a novel function of ILC3s in limiting tissue damage by preserving tissue-specific stem cells.The intestinal epithelium combines efficient uptake of nutrients and water while providing a physical barrier between the intestinal microbiota and the body (Peterson and Artis, 2014). Damage sustained by intestinal epithelial cells (IECs) needs to be swiftly and efficiently repaired to prevent inappropriate immune responses to commensal bacteria. Intestinal damage is an early event in the development of both graft-versus-host disease (Reddy and Ferrara, 2003) and alimentary mucositis (Sonis, 2004) and a driver of bacterial translocation and T cell activation in inflammatory bowel disease (Salim and Söderholm, 2011).A major pathway involved in the intestinal epithelial response to damage is the activation of Stat3, which is expressed along the crypt–villus axis of the intestinal epithelium (Grivennikov et al., 2009; Heneghan et al., 2013). Phosphorylated Stat3 translocates to the nucleus and activates genes involved in proliferation, survival, and mucosal defense (Bollrath et al., 2009; Pickert et al., 2009; Ernst et al., 2014). Mutations in STAT3 have been identified as susceptibility factors for inflammatory bowel disease (Bollrath et al., 2009; Anderson et al., 2011; Demaria et al., 2012), and in mice, upon DSS-induced colitis, epithelial Stat3 is required for mucosal wound healing (Pickert et al., 2009).Intestinal regeneration depends on the continuous differentiation of epithelial cells from crypt-residing intestinal stem cells (ISCs; Potten et al., 1978; Günther et al., 2013; Ritsma et al., 2014). Even though multiple intestinal progenitor cells have been described, the best-characterized populations are the Lgr5-expressing cells that reside at the crypt bottom, interspersed with Paneth cells. These stem cells have the ability to give rise to all IECs ex vivo (Sato et al., 2009). Similar to its role in differentiated epithelial cells, Stat3 activation is also an important pathway for survival of intestinal epithelial stem cells (Matthews et al., 2011).Type 3 innate lymphoid cells (ILC3s) are innate immune cells that reside in the lamina propria of both the small and large intestines and are involved in tissue homeostasis, early defense against enteric pathogens, and containment of microbiota (Spits and Cupedo, 2012; Artis and Spits, 2015). In the intestines, multiple ILC3 subsets exist, two of which can be distinguished by mutual exclusive expression of the natural cytotoxicity receptor NKp46 and the chemokine receptor CCR6 (Sawa et al., 2010; Reynders et al., 2011). Most Nkp46⁺ ILC3s are found dispersed throughout the lamina propria, a localization that depends on the expression of CXCR6 (Satoh-Takayama et al., 2014). In contrast, the majority of CCR6⁺ ILC3s are located in close proximity to the intestinal crypts in anatomically defined sites known as cryptopatches (Kanamori et al., 1996). Recent findings indicated that under inflammatory conditions, such as experimental graft-versus-host disease, ILC3s can interact with the epithelial stem cells in the crypts, protecting them from T cell–mediated killing (Hanash et al., 2012).The well-known ability of ILC3s to condition the local microenvironment, the close proximity of ILC3s to intestinal crypts, and the ability of ILC3s to communicate with epithelial stem cells led us to hypothesize that ILC3s are involved in directing intestinal epithelial responses to tissue damage. Using the methotrexate (MTX) model of small intestinal damage, we now show that ILC3s are activated immediately after MTX administration, leading to a rapid activation of epithelial Stat3 and maintenance of ISCs. Our data reveal a novel function for ILC3s as organizers of the intestinal epithelial response to tissue damage through activation of epithelial cells and maintenance of ISCs and suggest that ILC3s might in future be therapeutically harnessed to prevent stem cell loss during chemotherapy.